Secondary Metabolites:- These are organic compounds produced by any lifeform, e.g. bacteria, fungi, animals, or plants, which are not directly involved in the metabolism for normal growth, development, or reproduction of the organism.
Classification:-
1. Terpenes
2. Phenolics
3. Alkaloids
Intellectual Property Rights (IPRs):- These are the legal rights granted to individuals or organizations to protect their intellectual creations, such as inventions, literary and artistic works, designs, symbols, names, and trademarks. These rights give the creator exclusive control over the use, production, and commercialization of their creations for a specified period.
PCR:- The process of multiplication of DNA segments using DNA polymerase and DNA primers is called PCR.
• Discovery:- The PCR technique was discovered by Kary Mullis in 1985.
Gene tagging:- It is a molecular biology technique used to identify, isolate, and locate a gene of interest by using a known DNA sequence or genetic marker that is closely linked to the target gene. The marker acts as a "tag," making it easier to track the inheritance and position of the gene.
.png)
.png)
Southern blotting:-
- Discovered by Edward M. Southern at Edinburgh University in 1970.
- It is designed to locate a particular sequence of DNA within a complex mixture. For example, it could be used to locate a particular gene within an entire genome.
Procedure:-
a. Digestion:- Digest the DNA with an appropriate restriction enzyme.
b. Electrophoresis:- Run the digest on an agarose gel.
c. Denaturation:- Denature the DNA on the gel. For this soak it in about NaOH, which would separate double-stranded DNA into single-stranded DNA. Only ssDNA can transfer.
d. Depurination:- Fragments greater than 15 kb are hard to transfer to the blotting membrane. Depurination with HCl takes the purines out, cutting the DNA into smaller fragments. Neutralize the acid after this step.
e. Blotting:-
- It is the transfer the denatured DNA to the membrane.
- A nitrocellulose membrane or nylon membrane is used. Nitrocellulose typically has a binding capacity of about 100µg/cm, while nylon has a binding capacity of about 500 µg/cm.
- Many scientists feel nylon is better since it binds more and is less fragile.
- Transfer is usually done by capillary action, which takes several hours. Capillary action transfer draws the buffer up by capillary action through the gel an into the membrane, which will bind ssDNA.
- You may use a vacuum blot apparatus instead of capillary action. In this procedure, a vacuum sucks SSC (Saline Sodium Citrate) through the membrane. This works similarly to capillary action, except more SSC goes through the gel and membrane, so it is faster (about an hour). (SSC provides the high salt level that you need to transfer DNA.)
- After you transfer your DNA to the membrane, treat it with UV light. This cross links (via covalent bonds) the DNA to the membrane. (You can also bake nitrocellulose at about 80C for a couple of hours, but be aware that it is very combustible.)
f. Hybridization:-
- It is the process of annealing the probe to the DNA on the membrane due to complementarity.
- Probing is often done with:
i. 32P labeled ATP
ii. Biotin / Streptavidin
iii. Bioluminescent probe
g. Visualization:-
- Visualize your radioactively labeled target sequence.
- If you used a radiolabeled 32P probe, then you would visualize by autoradiograph.
- Biotin / Streptavidin detection is done by colorimetric methods.
- Bioluminescent visualization uses luminesence.
.png)
Male Sterility:- When the pollens of a plant are dead and inactive, while the female gametes are alive and active, it is called male sterility. This occurs very rarely. This occurs due to mutation. There are three types of male sterility: -
1. Genetic Male Sterility (GMS)
2. Cytoplasmic Male Sterility (CMS)
3. Cytoplasmic Genetic Male Sterility (CGMS)
1. Genetic Male Sterility (GMS):- When male sterility is controlled by a recessive gene present in the nucleus, it is called genetic male sterility (GMS).
MS gene = For male fertility
ms gene = For male sterility
MS MS = Male fertile plant
MS ms = Male fertile plant
ms ms = Male sterile plant
Maintenance:- For maintaining GMS, male sterile plants are cross heterozygous male fertile plants. As a result, 50 percent male sterile plants are obtained in each generation.
Application in hybrid seed production:- For hybrid seed production, the male sterile line is used as the female parent and crosses with the appropriate male parent line. Hybrid seeds are produced using GMS in Castor and Pigeon pea crops.
EGMS (Environment sensitive GMS):- The environment has a great impact on genetic male sterility. Genetic male sterility is determined by photo period and temperature. This property is used in hybrid seed production in paddy crop. Thus, there are two types of EGMS:
i. TGMS (Temperature sensitive GMS)
ii. PGMS (Photoperiod sensitive GMS)
i. TGMS (Temperature sensitive GMS):- Temperature has an effect on genetic male sterility in paddy crop. At temperatures below 28 ° C, paddy plants are completely male fertile while at temperatures above 30 °C, paddy plants are completely male sterile.
Maintenance of TGMS line:- For this, paddy plants are grown in an area where the day temperature remain below 28 ° C. At this temperature, paddy plants are fully male fertile. Now allow self-pollination in plants, resulting in the production of TGMS line seeds and then plants.
Hybrid Seed Production:- For this, paddy plants are grown in an area where the temperature is above 30 ° C. At this temperature paddy plants are fully male sterile which can be used as female parent. Now these male sterile female parent plants are crossed with the appropriate male parent plant, resulting in the production of hybrid seeds.
ii. PGMS (Photoperiod sensitive GMS):- Photo period also has an effect on genetic male sterility in paddy crop. Paddy plants are fully male fertile at 10 hours of light period, while at 14 hours of light, paddy plants are completely male sterile.
Maintenance of PGMS line:- For this, paddy plants are grown in an area where light period remain available for 10 hours. Paddy plants are fully male fertile in this condition. Now allow self-pollination in plants, resulting in the production of PGMS line seeds and then plants.
Hybrid Seed Production:- For this, paddy plants are grown in an area where the light period lasts for 14 hours. Paddy plants are fully male sterile in this condition which can be used as female parent. Now these male sterile female parent plants are crossed with the appropriate male parent plant, resulting in the production of hybrid seeds.
2. Cytoplasmic Male Sterility (CMS):- When male sterility is controlled by cytoplasm, it is called cytoplasmic male sterility (CMS). When fertilization occurs, only the nucleus comes from the male parent, while both the nucleus and cytoplasm come from the female parent. So when the male sterile female parent is crossed with the male parent, all the plants produced from the hybrid seeds are male sterile as shown in the diagram below.
Hybrid Seed Production:- CMS cannot be used for hybrid seed production in crops in which seed is of economic importance. But in crops where a somatic part is of economic importance rather than seed, CMS is used for hybrid seed production. Such as onion, garlic, carrot, radish etc.
3. Cytoplasmic Genetic Male Sterility (CGMS):- When male sterility is controlled by both nucleus and cytoplasm, it is called cytoplasmic genetic male sterility (CGMS). Cytoplasm is the main controller. The dominant gene present in the nucleus is called restorative gene, which acts to restore male fertility by preventing the male sterile effect of cytoplasm. We denote this as R letter. While r is the inactive gene.
R gene = For male fertility
r gene = For male sterility
RR = Male fertile plant
Rr = Male fertile plant
rr = Determined by cytoplasm.
Different combinations of nucleus and cytoplasm are given in the following diagram: -
Hybrid Seed Production:- CGMS is most commonly used in hybrid seed production. A three line system has been developed for hybrid seed production using CGMS which consists of the following three lines:
i. A – line:- It is a male sterile line that is used as a female parent.
ii. B – line:- It is a male fertile line which is used as a male parent. It is the maintainer line that works to maintain the male sterile A - line.
iii. R – line:- It is a male fertile line which is used as a male parent. It is a restore line that works to restore the male fertility in hybrid plants.
To maintain the A-line, its cross is made with the B-line. For hybrid seed production, the A-line crosses with the R-line as shown in the diagram below.
.png)
Transgenic Plant:-
• Definition:- Stable transformation is achieved when a desired gene is integrated into the genome of a plant, this plant is called transgenic plant.
• These transgenic plants are developed for the following purposes:-
1. Insect Resistance
2. Virus Resistance
3. Seed Protein Quality
4. Gene Silencing
5. Male Sterility
6. Biochemical Production
1. Insect Resistance:-
cry gene transfer:- The cry gene is found in the plasmid of Bacillus thuringiensis (Bt) bacteria. This cry gene makes Crystal protein which is insecticidal.
Ø The cry gene was isolated from the plasmid of Bt and integrated into the cotton genome to form a transgenic plant called Bt-cotton.
Ø This Bt - cotton is resistant to boll worm.
Ø Crystal protein perforates the larva's alimentary canal leading to its death.
2. Virus Resistance:-
cp – gene transfer:- The coat protein is transferred from the TMV virus to the tobacco plant.
Ø The RNA of TMV virus is not come out in the presence of cp-protein. Due to which the tobacco plant becomes resistant to mosaic disease.
Ø Other examples: - Tomato, Alfalfa, Beetroot, Potato
3. Seed Protein Quality:-
SFA8 gene transfer:- In sunflower seeds, the SFA8 gene produces a protein that contains an excess of sulfur-rich amino acids methionine and cysteine. While the proteins present in pea seeds lack methionine and cysteine amino acids.
Ø Therefore the SFA8 gene is transferred to pea seeds.
4. Gene Silencing:-
Slow ripening tomatoes:-
Ø PG (Poly Galacturonase) enzyme digest the pectin. Due to which fruits become soft and fluffy quickly and ripen quickly.
Ø Anti-sense genes are integrated into the genome of tomato plants against genes that code PG enzymes. This inhibits the expression of PG gene and the tomato fruit ripens late.
Ø Antisense gene made against PG gene inhibits its expression, which causes tomato to ripen late and can be easily exported.
5. Male Sterility:-
Producing male sterility:-
Ø Flavonoid are essential for maturation of pollens. If we stop its formation then male sterility can be created in the plant.
Ø The CHS (Chalcone Synthase) enzyme is required for flavonoid synthesis.
Ø Anti-sense genes are integrated into the plant's genome against genes that code CHS.
Ø Absence of a flavonoid causes pollen to become inactive. The plant becomes male sterile.
6. Biochemical Production:-
• PHB (Poly Hydroxy Butyrate):- It is prepared from Acetyl CoA. From this bio-degradable plastic is made.
• The transgenic plant is developed by isolating the phb - B and phb - C genes from the Alcaligenes eutrophus bacterium and transfer to Arabidopsis thaliana. Now PHB is obtained from this plant.
Golden Rice:-
• The genes that form β - carotene are integrated into the embryo of rice within the genome to prepare golden rice.
• 2 genes are isolated from Daffodils plant and 1 gene from Erwinia uredovora bacterium.
• β - carotene is yellow in color, which makes rice yellow.
• β - carotene is the precurssor of vitamin A.
Socio-Economic Aspects of Biotechnology:- Biotechnology is the application of living organisms, cells, or biological systems to develop useful products and technologies for agriculture, medicine, industry, and environmental management. It has significantly influenced socio-economic development by improving food security, healthcare, employment, industrial productivity, and environmental sustainability. However, it also raises ethical, legal, and social concerns.
Socio-Economic Aspects of Biotechnology:-
1. Improvement in Agriculture:-
> Development of high-yielding and disease-resistant crop varieties.
> Production of drought-, salinity-, and pest-resistant crops through genetic engineering.
> Reduced dependence on chemical pesticides and fertilizers.
> Improved nutritional quality of crops (e.g., Golden Rice enriched with Vitamin A).
> Increased farmers' income through higher productivity.
2. Healthcare Benefits:-
> Production of vaccines, antibiotics, insulin, growth hormones, and monoclonal antibodies.
> Early diagnosis of diseases using molecular diagnostic tools such as PCR and ELISA.
> Gene therapy and stem cell therapy for treating genetic disorders.
> Lower mortality rates and improved quality of life.
> Rapid development of vaccines during disease outbreaks.
3. Industrial Development:-
> Production of enzymes, biofuels, organic acids, and biodegradable plastics.
> Improved efficiency in food, textile, paper, leather, and pharmaceutical industries.
> Reduced production costs through microbial fermentation.
> Promotion of environmentally friendly industrial processes.
4. Employment Generation:-
> Creation of jobs in biotechnology research laboratories, pharmaceutical industries, agriculture, food processing, and environmental management.
> Increased demand for skilled professionals in molecular biology, bioinformatics, genetic engineering, and biotechnology.
> Growth of biotechnology startups and entrepreneurship opportunities.
5. Economic Growth:-
> Biotechnology contributes significantly to national GDP through agricultural exports, pharmaceuticals, and industrial biotechnology.
> Increased foreign investment in biotechnology research and development.
> Promotion of innovation and commercialization of new technologies.
> Strengthening of the bioeconomy.
6. Environmental Sustainability:-
> Bioremediation helps clean polluted soil and water.
> Biofertilizers and biopesticides reduce environmental pollution.
> Development of renewable biofuels decreases dependence on fossil fuels.
> Conservation of biodiversity through tissue culture and cryopreservation.
7. Food Security:-
> Increased food production to meet the needs of a growing population.
> Enhanced shelf life and nutritional quality of food products.
> Reduction in post-harvest losses through biotechnology-based preservation methods.
8. Ethical and Social Issues:-
> Concerns about the safety of genetically modified (GM) crops.
> Potential health and environmental risks.
> Ethical issues related to cloning, gene editing, and stem cell research.
> Public acceptance varies due to cultural and religious beliefs.
9. Intellectual Property Rights (IPR):-
> Biotechnology innovations are protected by patents.
> Patents encourage research and investment.
> High cost of patented technologies may limit access for poor farmers and developing countries.
> Need for balancing innovation with public welfare.
10. Challenges:-
> High research and development costs.
> Unequal access to biotechnology between developed and developing countries.
> Biosafety and biosecurity concerns.
> Need for proper regulations, public awareness, and skilled manpower.
.png)
.png)
Transcription in eukaryotes:-
Steps of transcription:-
i. Initiation:- Here, the enzyme RNA polymerase binds at the promoter site of DNA. This causes the local unwinding of the DNA double helix. An initiation factor (σ factor) present in RNA polymerase initiates the RNA synthesis.
ii. Elongation:- The RNA chain is synthesized in the 5’-3’ direction. In this process, activated ribonucleoside triphosphates (ATP, GTP, UTP & CTP) are added. This is complementary to the base sequence in the DNA template.
iii. Termination:- A termination factor (ρ factor) binds to the RNA polymerase and terminates the transcription.
In eukaryotes, there are 2 additional complexities-
i. There are 3 RNA polymerases:-
• RNA polymerase I:- Transcribes rRNAs (28S, 18S & 5.8S).
• RNA polymerase II:- Transcribes mRNA.
• RNA polymerase III:- Transcribes tRNA, 5S rRNA and snRNAs.
ii. The primary transcripts (hnRNA):- It contain both the exons and introns and is non-functional. Hence introns have to be removed. For this, it undergoes splicing process.
Heterogenous RNA Processing:- The hnRNA is the collective term for the unprocessed mRNA (pre-mRNA) molecules in the nucleus. It contain both the exons and introns and is non-functional. Hence introns have to be removed. For this, it undergoes the following processes-
i. Splicing:- From hnRNA introns are removed by the spliceosome and exons are joined together.
ii. Capping:- Here, a nucleotide methyl guanosine triphosphate (cap) is added to the 5’ end of hnRNA.
iii. Tailing (Polyadenylation):- Here, adenylate residues (200-300) are added at 3’-end. It is the fully processed hnRNA, now called mRNA.
Translation in Eukaryotes:- It takes place at ribosomes. Includes 4 steps-
1. Charging of tRNA (aminoacylation of tRNA):-
· Formation of peptide bond requires energy obtained from ATP.
· For this, amino acids are activated (amino acid + ATP) and linked to their cognate tRNA in the presence of aminoacyl tRNA synthetase. So the tRNA becomes charged.
2. Initiation:-
· It begins at the 5’-end of mRNA in the presence of an initiation factor.
· The mRNA binds to the small subunit of ribosome. Now the large subunit binds to the small subunit to complete the initiation complex.
· Large subunit has 2 binding sites for tRNA- aminoacyl tRNA binding site (A site) and peptidyl site (P site).
· Initiation codon for methionine is AUG. So methionyl tRNA complex would have UAC at the Anticodon site.
3. Elongation:-
· At the P site the first codon of mRNA binds with anticodon of methionyl tRNA complex.
· Another aminoacyl tRNA complex with an appropriate amino acid enters the ribosome and attaches to A site. Its anticodon binds to the second codon on the mRNA and a peptide bond is formed between first and second amino acids in presence of an enzyme, peptidyl transferase.
· First amino acid and its tRNA are broken. This tRNA is removed from P site & send to E site and second tRNA at the A site is pulled to P site along with mRNA. This is called translocation.
· Then 3rd codon comes on A site and a suitable tRNA with 3rd amino acid binds at the A site. This process is repeated.
· A group of ribosomes associated with a single mRNA for translation is called a polyribosome (polysomes).
4. Termination:-
· When aminoacyl tRNA reaches the termination codon like UAA, UAG & UGA, the termination of translation occurs. The polypeptide and tRNA are released from the ribosomes.
· The ribosome dissociates into large and small subunits at the end of protein synthesis.
· An mRNA has additional sequences that are not translated (untranslated regions or UTR). UTRs are present at both 5’-end (before start codon) and 3’-end (after stop codon). They are required for efficient translation process.
.png)
Nitrogen fixing genes and their genetic manipulation:-
Nif genes:-
1. Introduction:-
> Nif means:-
Ni = Nitrogen
f = fixation
> These are genes encoding enzymes involved in the fixation of atmospheric nitrogen.
> Nif genes also encode a number of regulatory proteins involved in nitrogen fixation.
> The nif genes are found in both free-living and symbiotic nitrogen-fixing bacteria.
> The primary enzyme encoded by the nif genes is the nitrogenase.
2. Regulation of Nif genes:- In most bacteria, regulation is done by NifA protein.
i. When there is not enough fixed nitrogen, NtrC triggers NifA expression, and NifA activates the nif genes.
ii. When there is not enough fixed nitrogen, NifL inhibit NifA expression, and nif genes remain inactive.
Reversible ADP-ribosylation:- It is an additional regulation mechanism found in Rhodospirillum rubrum. Reversible ADP-ribosylation of a specific arginine residue in the nitrogenase complex. When reduced nitrogen is present, DraG and DraT catalyze the ribosylation of arginine residue in the nitrogenase. It causes a barrier in the electron transfer flow and thereby inactivates nitrogenase activity.
3. Expression of Nif genes:-
> There are total 20 nif genes.
> nifH, nifD, and nifK:- They encode the nitrogenase subunits.
> nifE, nifN, nifU, nifS, nifV, nifW, nifX, nifB, and nifQ:- They encode proteins involved the assembly and incorporation of Fe and Mo atoms into the nitrogenase subunits.
> nifF and nifJ:- They encode proteins related to electron transfer taking place in the reduction process.
> nifA and nifL:- They are regulatory proteins in charge of regulating the expression of the other nif genes.
Nitrogen fixation:- It is a chemical process by which molecular nitrogen found in the air is converted into ammonia or related nitrogenous compounds.
1. Types of nitrogen fixation:-
a. Physical Nitrogen Fixation
b. Biological Nitrogen Fixation
a. Physical Nitrogen Fixation:-
i. Natural Nitrogen Fixation:- Under the influence of lightning and thunder, N2 and O2 of the air react to form nitric oxide (NO). The nitric oxides are again oxidized with oxygen to form nitrogen peroxide (NO2).
ii. Industrial Nitrogen Fixation:- Ammonia is produced industrially by direct combination of nitrogen with hydrogen (obtained from water) at high temperature and pressure. Later, it is converted into various kinds of fertilizers, such as urea etc.
b. Biological Nitrogen Fixation:- The conversion of atmospheric nitrogen into the nitrogenous compounds by living organisms is called biological nitrogen fixation. Only prokaryotes can fix nitrogen. Nitrogen fixation require anaerobic conditions because oxygen inactivates nitrogenase enzyme.
Hence for obligate anaerobes nitrogen fixation is easy, but in case of facultative anaerobes the nitrogen fixation occurs only in anaerobic conditions. In case of obligate aerobes the oxygen level inside the cell must be kept low for nitrogen fixation.
2. Nitrogen Fixers (Diazotrophs):- Among the earth’s organisms, only some prokaryotes like bacteria and cyanobacteria can fix atmosphere nitrogen. They are called nitrogen fixers or diazotrophs. They fix about 95% of the total global nitrogen fixed annually by natural process.
a. Asymbionts (Free living)
b. Symbionts
a. Asymbionts (Free living):-
i. Bacteria:- They add up to 10-25 kg, of nitrogen/ha/annum.
> Azotobacter (Aerobic, Saprophytic)
> Beijerinckia (Aerobic, Saprophytic)
> Clostridium (Anaerobic, Saprophytic)
> Desulphovibrio (Chemotrophic)
> Rhodopseudomonas (Photoautotrophic)
> Rhodospirillum (Photoautotrophic)
> Chromatium (Photoautotrophic)
ii. Blue Green Algae (Cyanobacteria):- Heterocysts are the special cells that fix nitrogen. They add 20-30 kg Nitrogen/ha/annum.
> Nostoc
> Anabaena
> Aulosira:- A. fertilissima is the most active nitrogen fixer in Rice fields.
> Cylindrospermum:- It is active in sugarcane and maize fields.
> Trichodesmium
b. Symbionts:- Live in close symbiotic association with other plants.
i. Blue Green Algae (Cyanobacteria):-
> Nostoc and Anabaena:- They are common symbionts in lichens, Anthoceros, Azolla and cycad roots.
> Anabaena azollae:- It is found in fronds of Azolla pinnata (a water fern). It is often inoculated to Rice fields for nitrogen fixation.
ii. Bacteria:-
> Rhizobium:- It is aerobic, gram negative nitrogen fixing bacterial symbionts of legume roots. Sesbania rostrata has Rhizobium in root nodules and Aerorhizobium in stem nodules.
> Frankia:- It is symbiont in root nodules of many non-leguminous plants like Casuarina and Alnus.
> Xanthomonas and Mycobacterium:- They occur as symbiont in the leaves of some members of the families Rubiaceae and Myrsinaceae (e.g., Ardisia).
3. Rhizobium Nitrogen Fixation:-
> Rhizobium bacteria:-
i. Free living
ii. Gram negative
iii. Aerobic
iv. Soil bacteria
> Rhizobium becomes anaerobic upon entry into roots.
> Leghaemoglobin (legHb or symbiotic Hb):-
- It is a pink coloured pigment.
- It occurs in the root nodules of leguminous plants.
- It acts as an oxygen scavenger. It provides anaerobic conditions for the nitrogenase enzyme and protects the enzyme from inactivation.
> Two main steps:-
a. Nodule formation
b. Nitrogen fixation
a. Nodule formation:- Root nodule formation is initiated, when the soil contains a low level of nitrogen. Steps of nodulation are:
i. Aggregation:- Roots of legumes secrete flavonoids, which attracts rhizobia towards the root. Rhizobia aggregate around root hairs.
ii. Developmental changes:- Rhizobia secrete nod factors, which causes stimulate many developmental changes:
- Membrane depolarization
- Curling of root hairs
- Cell division in the root cortex
- Intracellular calcium movement
iii. Infection thread:- The nod factor attaches to receptors present on the plasma membrane of the root hairs, which leads to the formation of the infection thread.
iv. Entry:- Infection thread provides the passage to bacteria to enter epidermal cells. Rhizobia then enter cortex cells, each bacterium gets surrounded by a plant-derived membrane known as symbiosome.
v. Nodulation:- Nodule formation is initiated by chemicals produced by rhizobia. It is a result of calcium dependent signal transduction pathway, which triggers biochemical changes leading to cell division and nodule formation. Cytokinin also plays an important role in nodules formation.
vi. Bacteroids:- Within nodules, bacteria get differentiated into bacteroids, which fix nitrogen. The Rhizobia stop dividing, loose cell wall and become nitrogen fixing cells as led bacteroids . Vascular tissues are developed for nodules for exchange of nutrients.
b. Nitrogen fixation:-
- The nodule serves as site for N2 fixation.
- Nodule contains nitrogenase and leghaemoglobin.
- The nitrogenase has 2 components:
i. Molybdoferredoxin (Mo-Fe protein)
ii. Azoferredoxin (Fe-protein).
- The free di-nitrogen first bound to MoFe protein and is not released until completely reduced to ammonia.
- In this process ferredoxin serves as an electron donor to Fe-protein (nitrogenase reductase) which in turn hydrolyzes ATP and reduce MoFe protein, the MoFe protein in Turn reduce the substrate N2. The electrons and ATP are provided by photosynthesis and respiration of the host cells.
- Many intermediates are formed to form ammonia (NH3).
Dinitrogen → Hydrazine → Diamine → Ammonia
- Ammonia (NH3) is immediately protonated at physiological pH to form ammonium ion (NH4+). As NH4+ is toxic to plants, it is rapidly used near the site of generation to synthesize amino acids.
Genetic manipulation of nitrogen fixation:-
> The subunits of nitrogenase from different nitrogen fixing microorganisms can be mixed to produce
functional system
> Genes can be manipulated to improve the fixation of dinitrogen.
> The microorganisms are modified in the host so that they are unable to assimilate the fixed nitrogen until the function of the nitrogenase is over and release NH4+ directly to the plants.
> The nitrogen fixation can be regulated by leguminous plants by:-
i. Reducing the number of root nodule formation
ii. Regulating the carbon flow to the microorganisms.
> The well known nitrogen fixing bacteria K. pneumoniae resembles closely to non-nitrogen fixing bacterium E.coli. The genes of these two species could be transferred and expressed in either of the organisms.
- Nif- mutants of K. pneumoniae that are deficient in fixing nitrogen are located between genes for histidine biosynthesis (his) and shikimik acid uptake (shi A).
- The his and nif regions can be actively transferred from a strain of K. pneumoniae to an E.coli
strain which require histidine.
- E.coli cells that do not require histidine anymore have acquired the ability to fix nitrogen.
- The conjugative plasmid pRDI that picked up the nif and his genes was selected and transferred to other bacterial genome.
- The gene, nifL, that serves as the repressor of nitrogen fixation can be deleted thus allowing constitutive expression of nif promoter mediated by nifA and ntrC (nitrogen regulator) gene products.
- The nitrogenase activity has been observed in E.coli that carries the nif plasmid pRDI which was very
much similar to that of K. pneumoniae strain.
> Agrobacterium tumefaciens, an obligate aerobe not resembling Klebsiella did not result in nitrogen fixing recombinants upon transfer of pRDI to itself.
> Mutants of Azotobacter vinelandii that lack either of the components of nitrogenase regained the
nitrogenase activity when pRDI was transferred to them.
> Nitrogenase can be protected from the inhibitory action of oxygen by a protein leghaemoglobin. The genes encoding these proteins can be isolated and transferred to other nitrogen fixing systems so as to
protect the nitrogenase from oxygen activity.
> Nitrogenase activity is correlated with the hydrogenase activity that evolve hydrogen hence it
requires more energy. The energy can be saved if the evolved hydrogen is further reduced to water
releasing electrons. Many nitrogen fixing bacteria possess ‘uptake hydrogenases’ which consists of two subunits HupS and HupL and it is advantageous to introduce it together with the nif genes into hosts that do not possess uptake hydrogenase system.
.png)
Intellectual Property (IP):- IP refers to creations of the human mind such as inventions, literary and artistic works, designs, symbols, names, images, software, and new plant varieties that have commercial or economic value. Intellectual Property Rights (IPRs) provide legal protection to creators and inventors by giving them exclusive rights over the use of their creations for a specified period.
Rights Available to Protect Intellectual Property (IPRs):-
1. Patent:-
> Protects new inventions, products, and processes.
> Gives the inventor exclusive rights to make, use, sell, or license the invention.
> Generally valid for 20 years from the filing date.
> Example: A genetically engineered crop or a new pharmaceutical drug.
2. Copyright:-
> Protects original literary, artistic, musical, and software works.
> Prevents unauthorized copying, reproduction, or distribution.
> Protection usually lasts for the author's lifetime plus 60 years in India.
> Example: Books, research papers, computer software.
3. Trademark:-
> Protects brand names, logos, symbols, slogans, and product names.
> Helps distinguish one company's goods or services from another's.
> Renewable every 10 years.
> Example: Company logos and brand names.
4. Industrial Design:-
> Protects the aesthetic or ornamental appearance of a product.
> Covers shape, pattern, configuration, or decoration.
> Example: Packaging design of a biotechnology product.
5. Geographical Indication (GI):-
> Protects products that originate from a specific geographical region and possess unique qualities due to that origin.
> Example: Darjeeling Tea, Basmati Rice.
6. Plant Breeders' Rights (PBR):-
> Protects newly developed plant varieties.
> Grants breeders exclusive rights to produce, market, and sell seeds.
> In India, protected under the Protection of Plant Varieties and Farmers' Rights (PPV&FR) Act, 2001.
7. Trade Secret:-
> Protects confidential business information, formulas, manufacturing methods, and processes.
> Protection continues as long as the information remains secret.
> Example: Industrial fermentation techniques.
.png){kind=small}
.png){kind=small}
.png){kind=small}
.png){kind=small}
.png){kind=small}
.png){kind=small}
.png){kind=small}
.png){kind=small}
.png){kind=small}
.bmp)
.bmp)
.bmp)
.bmp)
.bmp)
.bmp)
.bmp)
.bmp)
.png){kind=small}
.png){kind=small}
.png)
.bmp)
.bmp)
.bmp)
.png){kind=small}
.png){kind=small}